Increases in impervious surfaces caused by urbanization has led to higher volumes and rates of stormwater runoff that transports urban pollutants directly into natural waterways. Bioretention cells (BRCs) are vegetated soil systems designed to intercept stormwater runoff and reduce loads of water and contaminants discharged to surface waters. Nitrogen removal efficiency is highly variable and improvements are constrained by a poor understanding of the physical, biological, and chemical processes that occur within a BRC. The objectives of this study are to characterize and quantify the microbial communities in a range of existing BRCs, and determine which design factors have the greatest impact on denitrification, a microbial process responsible for removing nitrogen from stormwater. We sampled 23 BRCs throughout MD, VA, and NC, and quantified patterns in populations of denitrifying bacteria, denitrification potential, and microbial community structure within the soil medium. We found the greatest denitrifier populations and denitrification potential in the upper layer of the soil medium, which does not coincide with the internal water storage zone that is engineered to harbor anaerobic conditions favorable to denitrifying bacteria at the bottom of recent BRC designs. Results indicate that BRC vegetative cover, soil media nitrogen, and organic carbon concentrations are among the variables that facilitate nitrifying and denitrifying bacteria populations in BRCs. Bacterial community composition was most different between the top and bottom samples of the BRCs while fungal community composition differed most by BRC vegetative cover. Both fungal and bacterial community compositions were influenced by nitrogen and carbon concentrations.